- Email: [email protected]
Beneficial effects of prostglandin on rat cardiac allografts with cyclosporine immunosuppression
Beneficial Effects of Prostglandin on Rat Cardiac Allografts With Cyclosporine Immunosuppression Y.B. Koh, I.S. Moon, and S.C. Park ABSTRACT Graft rejection is characterized by cellular infiltration, reduction in blood flow, and intravascular coagulation, finally resulting in graft failure and absolute increase in thromboxane and relative decrease in prostacyclin synthesis. It is also suspected that prostaglandin itself could cause prolongation of allograft survival. In this experimental study, after successful heterotopic cardiac transplantation in rats, CyA was administered by intramuscular injection and prostaglandin E1 (PGE1) given into the peritoneal cavity. Each dose was according to the experimental designs. The authors found that PGE1 in addition to CyA could provide beneficial effects with significantly increased cardiac allograft survival, with amelioration of pathological changes in allograft rejection reaction.
A
LLOGRAFT REJECTION is characterized by cellular infiltration with reduction in blood flow and intravascular coagulation, finally resulting in transplant organ failure. In these events an absolute increase in thromboxane and a relative decrease in prostacyclin synthesis in the allograft is suspected to be the basic phenomenon in acute rejection.1–3 Recently, it has been suggested that the reduction in renal blood flow in cyclosporine (CyA)induced nephrotoxicity may be due to inhibition of endothelial production of prostacyclin (PGI2) by CyA.4 Several animal experiments have been reported that prostaglandin prolongs allograft survival.5–7 The present study was undertaken to investigate the effects of exogenous prostaglandin E1 (analogue of PGI2) upon rat cardiac allografts during CyA immunosuppression. MATERIALS AND METHODS Gunn rat hearts were transplanted into Wistar rats by anastomosing donor ascending aorta and pulmonary artery to recipient abdominal aorta and vena cava, respectively. Experimental rats were divided into four groups: group I (control, no treatment); group II (CyA 5.0 mg/kg/d, IM); group III (PGE1 1 g/kg/d, intraperitoneal); and group IV (CyA-PGE1). Each group consisted of 10 rats. The beat of cardiac allografts was palpated daily, cessation of heartbeat was defined as rejection. All rats were killed on posttransplant day 7, regardless of cardiac status. Cardiac allografts and spleens were excised for microscopic examination and for counts of splenic lymphocyte stimulation indices (SLSI) with phytohemagglutinin (PHA) and concanavalin A (ConA). Microscopic examination of excised hearts after hematoxylin-eosin (H-E) staining was done to compare each group. Pathological grading of vascular © 2005 by Elsevier Inc. All rights reserved. 360 Park Avenue South, New York, NY 10010-1710 Transplantation Proceedings, 37, 137–138 (2005)
thrombosis, interstitial edema, interstitial hemorrhage, mononuclear cell infiltration, and myocardiac necrosis were used to classify the reaction as severe, moderate, or mild. Splenic lymphocyte stimulation experiments were done by a modification of the method described by Miller and Creaghe.8 Splenic lymphocytes were obtained after centrifugation of splenocytes treated with hypaque solution and cultured in RPMI 1640, NaHCO3, and calf serum under the condition of 5% CO and 37°C. Then, mitogen (PHA or ConA) was added, and the lumphocytes were cultured again for 72 hours. Finally, H3-thymidine was added and cultured again for 16 hours; cpm were counted by -counter. Control cpm was estimated without mitogen stimulation and test cpm with mitogen. Each SLSI was calculated as the test cpm divided by control cpm.
RESULTS
Nine hearts of group I and two hearts of group III, stopped beating within 7 days; postoperative, all cardiac allografts of groups II and IV continued beating for the entire experimental period. Various pathological changes consistent with rejection were seen in group I. There were minimal changes in groups II and IV compared to groups I and III. Findings of vascular thromboses were mild in group IV compared to group II. Overall pathological findings of group III was less severe than group I (Table 1). SLSI of From the Kang Nam St. Mary’s Hospital, Catholic University Medical Center, Seoul, Republic of Korea. Address reprint requests to Dr Prof. Yong-Bok Koh, Kang Nam St Mary’s Hospital, Catholic University Medical Center, 505 Ban Po Dong, Seo Cho Ku, Seoul 137-040, Republic of Korea. E-mail: [email protected] 0041-1345/05/$–see front matter doi:10.1016/j.transproceed.2005.01.062 137
138
KOH, MOON, AND PARK Table 1. Pathological Grading of Microscopic Change Group I
Group II
Group III
Findings
Severe
Moderate
Mild
Severe
Moderate
Mild
Vascular thrombosis (n) Interstitial edema (n) Interstitial hemorrhage (n) Mononuclear cell infiltration (n) Myocardial necrosis (n)
7
2
1
1
7
2
7 6
2 3
1 1
1
2 3
8 6
6
4
2
5
5
4
3
1
groups II and IV were significantly lower than those of groups I and III (P ⬍ .05). There was no significant difference between groups II and IV. SLSI of group III were significantly lower than group I (P ⬎ .05; Table 2). DISCUSSION
This experimental study suggests that PGE1 administration in addition to CyA in rat cardiac allografts enhances CyA action, ameliorating the changes of vascular thrombosis. Possibly PGE1 can influence lymphocyte immunomodulation. Table 2. Lymphocyte Stimulation Indices
Group Group Group Group
I II III IV
*Only one rat was studied.
PHA (mean ⫾ SD)
ConA (mean ⫾ SD)
24.1* 10.8 ⫾ 3.1 18.3 ⫾ 3.4 9.2 ⫾ 3.2
36.1* 16.2 ⫾ 3.7 29.4 ⫾ 6.5 15.6 ⫾ 4.4
Severe
Group IV
Moderate
Mild
5
5
2 1
3 4
5 5
3
4
3
3
7
1
3
6
Severe
Moderate
Mild
3
7
1
2 3
8 6
1
4
5
3
7
REFERENCES 1. Foegh M, Winchester F, Zmudka M, et al: Murine i-T B2 in renal allograft rejection. Lancet 2:431, 1981 2. Coffman TM, Yarger E, Klotman PE: Functional role of thromboxane production by acutely rejecting renal allografts in rats. Clin Invest 75:1242, 1985 3. Tannenbaum S, Anderson CB, Sicard GA, et al: Prostaglandin synthesis associated with renal allograft rejection in the dog. Transplantation 37:438, 1984 4. Rosenthal RA, Chukwuogo NA, Ocasio V, et al: Cyclosporine inhibits endothelial cell prostacyclin production. Surg Res 46:593, 1989 5. Mundy AR: Prolongation of cat to dog renal xenograft survival with prostacyclin. Transplantation 30:226, 1980 6. Imura M, Higashi, Yada I, Namikawa S, et al: Effect of prostaglandin E1 on the prolongation of rat cardiac allograft survival. Transplant Proc 19(1 Pt 2):1312, 1987 7. Faist E, Schinkel C, Zimmer S: Update on the mechanisms of immune suppression of injury and immune modulation. World J Surg 20:454, 1996 8. Miller TE, Creaghe E: Quantitation of potential T-lymphocyte function in rats. Infect Immun 12:722, 1975
A
LLOGRAFT REJECTION is characterized by cellular infiltration with reduction in blood flow and intravascular coagulation, finally resulting in transplant organ failure. In these events an absolute increase in thromboxane and a relative decrease in prostacyclin synthesis in the allograft is suspected to be the basic phenomenon in acute rejection.1–3 Recently, it has been suggested that the reduction in renal blood flow in cyclosporine (CyA)induced nephrotoxicity may be due to inhibition of endothelial production of prostacyclin (PGI2) by CyA.4 Several animal experiments have been reported that prostaglandin prolongs allograft survival.5–7 The present study was undertaken to investigate the effects of exogenous prostaglandin E1 (analogue of PGI2) upon rat cardiac allografts during CyA immunosuppression. MATERIALS AND METHODS Gunn rat hearts were transplanted into Wistar rats by anastomosing donor ascending aorta and pulmonary artery to recipient abdominal aorta and vena cava, respectively. Experimental rats were divided into four groups: group I (control, no treatment); group II (CyA 5.0 mg/kg/d, IM); group III (PGE1 1 g/kg/d, intraperitoneal); and group IV (CyA-PGE1). Each group consisted of 10 rats. The beat of cardiac allografts was palpated daily, cessation of heartbeat was defined as rejection. All rats were killed on posttransplant day 7, regardless of cardiac status. Cardiac allografts and spleens were excised for microscopic examination and for counts of splenic lymphocyte stimulation indices (SLSI) with phytohemagglutinin (PHA) and concanavalin A (ConA). Microscopic examination of excised hearts after hematoxylin-eosin (H-E) staining was done to compare each group. Pathological grading of vascular © 2005 by Elsevier Inc. All rights reserved. 360 Park Avenue South, New York, NY 10010-1710 Transplantation Proceedings, 37, 137–138 (2005)
thrombosis, interstitial edema, interstitial hemorrhage, mononuclear cell infiltration, and myocardiac necrosis were used to classify the reaction as severe, moderate, or mild. Splenic lymphocyte stimulation experiments were done by a modification of the method described by Miller and Creaghe.8 Splenic lymphocytes were obtained after centrifugation of splenocytes treated with hypaque solution and cultured in RPMI 1640, NaHCO3, and calf serum under the condition of 5% CO and 37°C. Then, mitogen (PHA or ConA) was added, and the lumphocytes were cultured again for 72 hours. Finally, H3-thymidine was added and cultured again for 16 hours; cpm were counted by -counter. Control cpm was estimated without mitogen stimulation and test cpm with mitogen. Each SLSI was calculated as the test cpm divided by control cpm.
RESULTS
Nine hearts of group I and two hearts of group III, stopped beating within 7 days; postoperative, all cardiac allografts of groups II and IV continued beating for the entire experimental period. Various pathological changes consistent with rejection were seen in group I. There were minimal changes in groups II and IV compared to groups I and III. Findings of vascular thromboses were mild in group IV compared to group II. Overall pathological findings of group III was less severe than group I (Table 1). SLSI of From the Kang Nam St. Mary’s Hospital, Catholic University Medical Center, Seoul, Republic of Korea. Address reprint requests to Dr Prof. Yong-Bok Koh, Kang Nam St Mary’s Hospital, Catholic University Medical Center, 505 Ban Po Dong, Seo Cho Ku, Seoul 137-040, Republic of Korea. E-mail: [email protected] 0041-1345/05/$–see front matter doi:10.1016/j.transproceed.2005.01.062 137
138
KOH, MOON, AND PARK Table 1. Pathological Grading of Microscopic Change Group I
Group II
Group III
Findings
Severe
Moderate
Mild
Severe
Moderate
Mild
Vascular thrombosis (n) Interstitial edema (n) Interstitial hemorrhage (n) Mononuclear cell infiltration (n) Myocardial necrosis (n)
7
2
1
1
7
2
7 6
2 3
1 1
1
2 3
8 6
6
4
2
5
5
4
3
1
groups II and IV were significantly lower than those of groups I and III (P ⬍ .05). There was no significant difference between groups II and IV. SLSI of group III were significantly lower than group I (P ⬎ .05; Table 2). DISCUSSION
This experimental study suggests that PGE1 administration in addition to CyA in rat cardiac allografts enhances CyA action, ameliorating the changes of vascular thrombosis. Possibly PGE1 can influence lymphocyte immunomodulation. Table 2. Lymphocyte Stimulation Indices
Group Group Group Group
I II III IV
*Only one rat was studied.
PHA (mean ⫾ SD)
ConA (mean ⫾ SD)
24.1* 10.8 ⫾ 3.1 18.3 ⫾ 3.4 9.2 ⫾ 3.2
36.1* 16.2 ⫾ 3.7 29.4 ⫾ 6.5 15.6 ⫾ 4.4
Severe
Group IV
Moderate
Mild
5
5
2 1
3 4
5 5
3
4
3
3
7
1
3
6
Severe
Moderate
Mild
3
7
1
2 3
8 6
1
4
5
3
7
REFERENCES 1. Foegh M, Winchester F, Zmudka M, et al: Murine i-T B2 in renal allograft rejection. Lancet 2:431, 1981 2. Coffman TM, Yarger E, Klotman PE: Functional role of thromboxane production by acutely rejecting renal allografts in rats. Clin Invest 75:1242, 1985 3. Tannenbaum S, Anderson CB, Sicard GA, et al: Prostaglandin synthesis associated with renal allograft rejection in the dog. Transplantation 37:438, 1984 4. Rosenthal RA, Chukwuogo NA, Ocasio V, et al: Cyclosporine inhibits endothelial cell prostacyclin production. Surg Res 46:593, 1989 5. Mundy AR: Prolongation of cat to dog renal xenograft survival with prostacyclin. Transplantation 30:226, 1980 6. Imura M, Higashi, Yada I, Namikawa S, et al: Effect of prostaglandin E1 on the prolongation of rat cardiac allograft survival. Transplant Proc 19(1 Pt 2):1312, 1987 7. Faist E, Schinkel C, Zimmer S: Update on the mechanisms of immune suppression of injury and immune modulation. World J Surg 20:454, 1996 8. Miller TE, Creaghe E: Quantitation of potential T-lymphocyte function in rats. Infect Immun 12:722, 1975